NR AXVK
AU Roucou,X.; Goggin,K.; Beaudoin,S.; Grenier,C.; Brown,A.A.
TI PrP Aggresomes are Poly(A+)-Ribonucleoprotein Complexes that Induce a RNA-Dependent Protein Kinase(PKR)-Mediated Deficient Cell Stress Response
QU International Conference - Prion 2007 (26.-28.9.2007) Edinburgh International Conference Centre, Edinburgh, Scotland, UK - Book of Abstracts: Pathology and Pathogenesis P03.105
IA http://www.prion2007.com/pdf/Prion Book of Abstracts.pdf
PT Konferenz-Poster
AB
Background: The cause of neurodegeneration in prion diseases is not well understood, and there is much evidence that argues against the direct neurotoxicity of PrPsc. Some attention has recently turned toward exploring mistrafficking and accumulation of PrP in the cytoplasm. Recent studies indicate that prion-infected cells produce juxtanuclear cytoplasmic inclusions termed PrP aggresomes, and that such aggregates may be present in the brain of infected mice. The mechanism of toxicity of PrP aggresomes has not been fully investigated.
Objective: We have investigated if PrP aggresomes induce a cellular stress by determining if cells producing such aggregates exhibit a spontaneous stress response.
Methods: PrP aggresomes were produced by transiently transfecting neuronal cells with a construct encoding PrP without the N- and C-terminal signal peptides.
Results: We report that cytoplasmic PrP aggregates initiate a cell stress response by activating the autophosphorylation of the RNA-dependent protein kinase (PKR) at threonine 446. Activated PKR phosphorylates the translation initiation factor eiF2a on serine 51, resulting in protein synthesis shut-off. However, other components of the stress response, including the assembly of poly(A)+ RNA-containing stress granules (SGs) and the synthesis of heat shock protein 70, is repressed. A phosphomimetic mutant of eiF2a normally sufficient to induce the assembly of SGs, was unable to induce the formation of SGs in cells producing PrP aggresomes. These observations suggested that poly(A)+ RNA are trapped and unable to be recruited into SGs. We verified this hypothesis by in situ hybridization experiments with fluorescent oligo-dT probes and confocal analysis. We observed the clustering of poly(A)+ RNA within PrP aggresomes. Affinity chromatography on oligo(dT)-cellulose demonstrated that PrP aggresomes purify with poly(A)+ RNA, and are therefore poly(A)+ ribonucleoprotein complexes in vivo.
Discussion: We have demonstrated that PrP aggresomes are poly(A)+ ribonucleoprotein complexes inducing a spontaneous stress response characterized by the activation of PKR, the phosphorylation of eiF2a, and repression of protein translation. However, aggregation of poly(A)+ prevent a full stress response. These mechanisms may not be obligatory lethal per se, but would likely result in premature cell death in the context of an acute environmental stress that would be otherwise dealt with an adequate stress response
AD X. Roucou, K. Goggin, S. Beaudoin, C. Grenier, A.-A. Brown, University of Sherbrooke, QC Canada, Biochemistry, Canada
SP englisch
PO Schottland